The discipline of metabolic engineering was defined fifteen years ago as “the improvement of cellular activities by manipulations of enzymatic, transport, and regulatory functions of the cell with the use of recombinant DNA technology” (Bailey 1991). Since that time, the field has witnessed a number of success stories with respect to the development of highly productive organisms, especially microbes. Initially, metabolic engineering efforts were primarily focused on improving the productivity of naturally-occurring metabolites in the target organisms, as is consistent with Bailey's original definition. More recently, the field has expanded to encompass a number of examples of introducing new enzyme activities into a host cell in order to produce non-natural products (Nielsen 2001). Non-natural products are defined in this case as compounds that are foreign to the production organism. Thus, such compounds may still be found in other organisms (e.g., plant natural products), or they may be novel, structurally distinct from those known to exist in nature.
Significant efforts have gone into the development of microorganisms to produce non-natural products. Examples include polyketides, with anti-infective, anti-tumor, and cholesterol-lowering properties (Pfeifer et al. 2001; Pfeifer et al. 2003); and isoprenoids, a class of compounds with uses that range from pigments (Mijts and Schmidt-Dannert 2003) to the treatment of malaria (Ro et al. 2006). The development of a fermentation process for the production of 1,3-propanediol at titers that exceed 125 g/L illustrates that the use of microbial chemical factories extends well beyond the synthesis of human therapeutics (Nakamura and Whited 2003). The advent of tools such as directed evolution and advances in the ability to rationally engineer or re-engineer proteins with desired activities against specified substrates enables one to imagine the ability to produce proteins capable of transforming an enormous range of chemical compounds into novel products (Lippow and Tidor 2007; Jiang et al. 2008). Indeed, efforts in biocatalysis have resulted in the identification of many enzymes displaying novel activities, which themselves are ideal candidates for directed evolution to expand the substrate and product repertoire even further (Bommarius and Polizzi 2006). Assembling several such proteins—either native or evolved towards optimal activity against a particular substrate—into a functioning metabolic pathway can result in the microbial production of a non-natural product. It is now conceivable that the production of many compounds of commercial value, traditionally reserved for the synthetic organic chemist, can be achieved with microbial systems.
One of the limitations for productivity that often arise in the development of microbial chemical factories is low product yield. Typical approaches towards increasing yields in metabolic engineering involve deleting the genes that encode for competing activities; however, this is not feasible when the mutation might severely limit cell growth or be lethal.